Beans are one of the most important crops in Mexico, the annual average intake of common beans is approximately 22 kg/capita (Castellanos et al 1997); interestingly black and pinto beans are the most widely consumed. Furthermore, the inventors found it intriguing that the incidence of mammary cancer is significantly lower in states where women consume black beans in contrast with other states where other types of beans are usually ingested. In 1997, the average death rate of females older than 25 years due to breast cancer was 14.8 per 100,000 females whereas for counterparts living in states where black beans are frequently consumed, the rate was 8.2 (CONAPO, 2004). The inventors suspected that the more than 40% lower risk might be at least partially attributable to consumption of black beans. As occurs in other parts of the world, a higher incidence of breast cancer death occurs in post-menopause females older than 40 years. The highest incidence of deaths due to breast cancer is in geriatric women older than 65 years (42.4/100,000 females). See also Azevedo et al, 2003 (mice fed with black bean diets may have shown a lower incidence of CP-induced DNA damage, but note that the study did NOT use purified compounds or fractions from purifying black bean samples, and employed a comet assay that only detects primary DNA lesions, which might or might not be converted into mutations, thus providing nothing more to the art than the anecdotal observations in CONAPO, 2004).
Several phenolic compounds have been reported from Phaseolus vulgaris, most of them phytoalexins isolated from fungal infected beans (Burden 1972, Perrin 1972, Kim, 1988, Beninger et al. 1998, Beninger et al. 1999). Likewise, other phenolics with potential nutraceutical properties have also been extracted from healthy black beans. Tannins may contribute at least 4% of the composition of the hulls and this percentage increases according to variety and/or storage conditions. Cardador-Martinez et al (2002) found that most of the phenolics and antioxidants in common beans were concentrated in the hulls or testas and that these antioxidants had free radical scavenging activity with antimutagenic activity. Different flavonoids are involved in the seed color of beans, in particular anthocyanins, which may account for 2.5% of the seed coat. The level of anthocyanins in black beans (at least 200 mg/100 g beans) is comparable to that reported in fruits such as berries (Takeoka et al 1997). Compounds that may have been previously identified have not been shown to have the utilities set forth herein.
Anthocyanins like other flavonoids seem to play an important role in the prevention of human diseases associated with oxidative stress. These properties have been attributed to their high antioxidant activity that ranges from 6-42% of the radicals scavenged using the DPPH (1,1-diphenyl-2-picrylhydrazyl) method, a value that is greatly affected by the presence of sugars bound to the molecules. Bound sugars diminish the antioxidant activity of flavonoids (Kähkönen and Heinonen, 2003). The anthocyanin literature does not disclose or suggest black bean and/or hull thereof extracts, compounds thereof, and uses thereof as herein set forth.
Another group of flavonoids previously reported in the literature are the isoflavones, which possess estrogenic and other biological activities. These compounds are considered to be non-nutritive; however, interest in these compounds has arisen because of their beneficial or nutraceutical effects (Setchel and Cassidy 1999). Tabor, U.S. Pat. No. 6,482,448 and Kelly, U.S. Pat. No. 6,497,906, involve formulations or supplements obtained from soybeans containing isoflavones daidzein, genistein, formononetin, biochanin A and glycitein, in different ratios and concentrations, to treat or prevent premenopausal symptoms, heart/cardiovascular related conditions, osteoporosis, breast/prostate cancers, endometrium abnormalities and head/brain symptoms including Alzheimer's Disease. There is no teaching or suggestion in such literature and patents of extracts from black beans and/or hulls thereof, compounds thereof, and the utilities therefore as herein disclosed.
Many natural compounds, such as flavonoids and saponins, occur mainly in their glycoside forms; the basic structure is substituted with at least one molecule of simple sugars such as glucose, galactose, arabinose, rhamnose and xylose. Removal of the bound sugars via fermentation or glycosidic enzymes yields extracts with higher bioactivity because the resulting aglycones have more affinity for cell receptors. Several U.S. Patents involve methods for preparing aglycone isoflavones enriched products (U.S. Pat. Nos. 6,579,561, 6,500,965, 6,146,668, 5,320,949; 5,352,384; 5,637,561 and 5,637,562). Izumi et al (2000) investigated the difference in the absorption of soy isoflavones aglycones and glycosides in humans and found that a higher plasma concentration was observed after aglycone intake (more than two times greater) than the levels observed after glucoside ingestion. Setchell et al (2001) determined that aglycones genistein and daidzein attained peak plasma concentration faster than their corresponding glycosides. U.S. Pat. No. 6,607,757 involves a soybean (Glycine max) extract having isoflavones and saponins to treat postmenopausal symptoms and breast and prostate cancer. Hendler et al., U.S. Pat. No. 6,541,613 involves a modification by esterification of isoflavones to promote bioavailability and enhance hydrosolubility. The esterified isoflavones may be employed therapeutically or prophylactically for a variety of conditions. These articles and patents do not disclose or suggest black bean and/or hull thereof extracts, compounds therefrom, and uses thereof as herein disclosed.
Thurn and Huang, U.S. Pat. No. 6,004,558 involves the preparation of therapeutic compositions comprising extracts of red clover (Tribolium sp) or soybeans from which the isoflavones were removed and nevertheless the therapeutic use of these extracts were effective to treat or prevent a variety of cancers. Therefore other types of flavonoids and/or phenolic compounds and/or triterpenes and/or other natural compounds are also useful to treat cancer. For example, Prochaska et al (U.S. Pat. No. 5,336,685) involves a method of inhibiting the growth of multidrug resistant cancer cells with flavonoids such as alpha and beta naphthoflavones, flavone, and 2,3 dihydroflavone. Buchholz et al., U.S. Pat. No. 6,514,527, involves a composition containing a mixture of bioflavonols isoqueracetin, queracetin 4-glycoside, rutin and quercetin possessing antioxidant and preventive properties against damage to human tissues and cardiovascular disease. On the other hand, Romancyzk et al., U.S. Pat. Nos. 6,562,863 and 6,479,539 pertains to cocoa (Theobroma cacao) extracts rich in polyphenols or procyanidins for use as antioxidant and antineoplastic agents. Recently, Bawadi et al (2005) demostrated that condensed tannins isolated from black beans did not interfere with the proliferation of normal human fibroblast lung cells, but inhibited the growth of Caco-2 colon, MCF-7 and Hs578T breast, and DU 145 prostatic cancer cells by disrupting the cells. Interestingly, they found that ATP levels were reduced in tannin-treated cancer cells, which implies reduced cell proliferation and migration activity, and gross morphological examination of tannin-treated cells suggested that cell death occurred by apoptosis. Morre et al (U.S. Pat. No. 6,410,061) involves extracts based on catechins obtained from green tea (Camellia sinensis) to treat cancers or solid tumors. Composition of catechins includes epigallocatechin gallate, epicatechin, epicatechin gallate and epigallocatechin. Epigallocatechin gallate, the major catechin found in green tea, blocks DNA transcription of a number of genes in cancer cell lines and therefore acted as anti-carcinogenic. These patents do not disclose or suggest black bean and/or hull thereof extracts, compounds therefrom, and uses thereof as herein disclosed.
As mentioned, most phenolic compounds found in black beans are concentrated in the testa (Cardador-Martinez et al, 2002). Sosulski and Dabrowski (1984) found that dehulling substantially reduced phenolic composition of pigeon pea, faba bean, mungo bean and lentils but had little effect on the phenolic composition of field pea, navy bean, lima bean or chickpea. Ronzio et al., U.S. Pat. No. 5,762,936 involves the preparation of extracts of seed coats of lentil (Lens esculenta) rich in condensed tannins, flavanones, flavanols, and phenolic acids that have the ability to quench free radicals and inhibit certain cells responsible for inflammation. The present inventors found no literature providing bioactivity in black bean hulls. But Grabiel et al. in a recently published U.S. Patent Application 20040131749A1 describes the potential use of phytochemicals, in particular polyphenolics extracted from beans that naturally are rich in anthocyanins, flavonols, proanthocyanidins, isoflavones, saponins, sapogenins, lectines, vitamins, minerals and functional proteins. They propose a method where edible beans and an aqueous extract are obtained, this last one a potential significant source of flavonols and anthocyanins that can be separated and used for treatment or reducing the probability of developing cancer, stroke, elevated blood cholesterol, hypertension, myocardial infarction, diabetes, obesity and inflammatory disorders in humans. These documents do not disclose or suggest (or disclose or suggest after the claim for priority of this invention, e.g. Bawadi et al. (2005)) black bean and/or hull thereof extracts, compounds therefrom, and uses thereof as herein disclosed; and, do not disclose or suggest the method for dehulling of the instant invention or the use of hulls as herein disclosed.
Thus, none of the scientific reports or patents sets forth or claims the production and utilization of black bean and/or hull thereof extracts, compounds therefrom, and the uses thereof disclosed herein, or the use of germination to increase the bioactivity of the compounds in black bean and/or hulls thereof as herein disclosed. Nor does the art particularly teach toward the use of black bean and/or hull extracts as the generous source of especially useful compounds which the inventors discovered in the present invention.